Exome and genome sequencing in adults with undiagnosed disease: a prospective cohort study.

BACKGROUND: Exome and genome sequencing have been demonstrated to increase diagnostic yield in paediatric populations, improving treatment options and providing risk information for relatives. There are limited studies examining the clinical utility of these tests in adults, who currently have limited access to this technology. METHODS: Patients from adult and cancer genetics clinics across Toronto, Ontario, Canada were recruited into a prospective cohort study evaluating the diagnostic utility of exome and genome sequencing in adults. Eligible patients were ≥18 years of age and suspected of having a hereditary disorder but had received previous uninformative genetic test results. In total, we examined the diagnostic utility of exome and genome sequencing in 47 probands and 34 of their relatives who consented to participate and underwent exome or genome sequencing. RESULTS: Overall, 17% (8/47) of probands had a pathogenic or likely pathogenic variant identified in a gene associated with their primary indication for testing. The diagnostic yield for patients with a cancer history was similar to the yield for patients with a non-cancer history (4/18 (22%) vs 4/29 (14%)). An additional 24 probands (51%) had an inconclusive result. Secondary findings were identified in 10 patients (21%); three had medically actionable results. CONCLUSIONS: This study lends evidence to the diagnostic utility of exome or genome sequencing in an undiagnosed adult population. The significant increase in diagnostic yield warrants the use of this technology. The identification and communication of secondary findings may provide added value when using this testing modality as a first-line test.

Homozygous GLUL deletion is embryonically viable and leads to glutamine synthetase deficiency.

Glutamine synthetase (GS) is the enzyme responsible for the biosynthesis of glutamine, providing the only source of endogenous glutamine necessary for several critical metabolic and developmental pathways. GS deficiency, caused by pathogenic variants in the glutamate-ammonia ligase (GLUL) gene, is a rare autosomal recessive inborn error of metabolism characterized by systemic glutamine deficiency, persistent moderate hyperammonemia, and clinically devastating seizures and multi-organ failure shortly after birth. The four cases reported thus far were caused by homozygous GLUL missense variants. We report a case of GS deficiency caused by homozygous GLUL gene deletion, diagnosed prenatally and likely representing the most severe end of the spectrum. We expand the known phenotype of this rare condition with novel dysmorphic, radiographic and neuropathologic features identified on post-mortem examination. The biallelic deletion identified in this case also included the RNASEL gene and was associated with immune dysfunction in the fetus. This case demonstrates that total absence of the GLUL gene in humans is viable beyond the embryonic period, despite the early embryonic lethality found in GLUL animal models.

Warsaw breakage syndrome: Further clinical and genetic delineation.

Warsaw breakage syndrome (WBS) is a recently recognized DDX11-related rare cohesinopathy, characterized by severe prenatal and postnatal growth restriction, microcephaly, developmental delay, cochlear anomalies, and sensorineural hearing loss. Only seven cases have been reported in the English literature, and thus the information on the phenotype and genotype of this interesting condition is limited. We provide clinical and molecular information on five additional unrelated patients carrying novel bi-allelic variants in the DDX11 gene, identified via whole exome sequencing. One of the variants was found to be a novel Saudi founder variant. All identified variants were classified as pathogenic or likely pathogenic except for one that was initially classified as a variant of unknown significance (VOUS) (p.Arg378Pro). Functional characterization of this VOUS using heterologous expression of wild type and mutant DDX11 revealed a marked effect on protein stability, thus confirming pathogenicity of this variant. The phenotypic data of the seven WBS reported patients were compared to our patients for further phenotypic delineation. Although all the reported patients had cochlear hypoplasia, one patient also had posterior labyrinthine anomaly. We conclude that while the cardinal clinical features in WBS (microcephaly, growth retardation, and cochlear anomalies) are almost universally present, the breakage phenotype is highly variable and can be absent in some cases. This report further expands the knowledge of the phenotypic and molecular features of WBS.

Genetic modifiers of abnormal organelle biogenesis in a Drosophila model of BLOC-1 deficiency.

Biogenesis of lysosome-related organelles complex 1 (BLOC-1) is a protein complex formed by the products of eight distinct genes. Loss-of-function mutations in two of these genes, DTNBP1 and BLOC1S3, cause Hermansky-Pudlak syndrome, a human disorder characterized by defective biogenesis of lysosome-related organelles. In addition, haplotype variants within the same two genes have been postulated to increase the risk of developing schizophrenia. However, the molecular function of BLOC-1 remains unknown. Here, we have generated a fly model of BLOC-1 deficiency. Mutant flies lacking the conserved Blos1 subunit displayed eye pigmentation defects due to abnormal pigment granules, which are lysosome-related organelles, as well as abnormal glutamatergic transmission and behavior. Epistatic analyses revealed that BLOC-1 function in pigment granule biogenesis requires the activities of BLOC-2 and a putative Rab guanine-nucleotide-exchange factor named Claret. The eye pigmentation phenotype was modified by misexpression of proteins involved in intracellular protein trafficking; in particular, the phenotype was partially ameliorated by Rab11 and strongly enhanced by the clathrin-disassembly factor, Auxilin. These observations validate Drosophila melanogaster as a powerful model for the study of BLOC-1 function and its interactions with modifier genes.

The white gene of Drosophila melanogaster encodes a protein with a role in courtship behavior.

The white gene of Drosophila melanogaster has been extensively studied, yet it is still not understood how its ectopic overexpression induces male-male courtship. To investigate the cellular basis of this behavior, we examined the sexual behavior of several classes of mutants. We find that male-male courtship is seen not only in flies overexpressing the white gene, but also in mutants expected to have mislocalized White protein. This finding confirms that mislocalizing White transporter in the cells in which it is normally expressed will produce male-male courtship behaviors; the courtship behavior is not an indirect consequence of aberrant physiological changes elsewhere in the body. Male-male courtship is also seen in some mutants with altered monoamine metabolism and deficits in learning and memory, but can be distinguished from that produced by White mislocalization by its reduced intensity and locomotor activity. Double mutants overexpressing white and with mutations in genes for serotonergic neurons suggest that male-male courtship produced by mislocalizing White may not be mediated exclusively by serotonergic neurons. We also find decreased olfactory learning in white mutants and in individuals with mutations in the genes for White's binding partners, brown and scarlet. Finally, in cultured Drosophila and mammalian cells, the White transporter is found in the endosomal compartment. The additional genes identified here as being involved in male-male courtship increase the repertoire of mutations available to study sexual behavior in Drosophila.